NOTES ON THE REARING OF ECHINOID LARViE. 95 



esculentus, but for the sake of comparison, cultures of the larvse of 

 Echinus miliaris were also prepared. 



The first condition for a healthy experiment is that only full-sized 

 and perfectly ripe individuals of both sexes should be used for fertilisa- 

 tion. It is generally possible to get a few eggs capable of fertilisation 

 from unripe individuals, but the resulting plutei are feeble, and soon 

 die. 



The fertilisation is best performed in a large square glass dish ; a 

 small quantity only of spermatozoa must be added to the clean sea- 

 water with which the dish is tilled. The ova are added in sufficient 

 quantity to make a single layer over the bottom. A piece of bolting- 

 silk of medium mesh will permit the ova to pass, but will keep back 

 pieces of ovary, and to a large extent unripe ova. 



The laboratory at Plymouth is provided with bolting-cloth of mesh 

 exactly suitable to the eggs of E. esculentus. The unripe eggs appear 

 to be surrounded by a thick, glassy membrane (entirely distinct from 

 the membrane formed after fertilisation), which prevents their passing 

 through the meshes of the silk. 



After fertilisation, the water full of spermatozoa must not be allowed 

 to stand more than twenty minutes before it is decanted off, and during 

 the first twenty-four hours the water covering the eggs should be 

 frequently changed. 



Next in importance to the proper choice of individuals for the 

 experiment is the selection of a proper source whence the sea-water is 

 to be obtained. Water obtained close inshore is perfectly useless. The 

 plutei will live for at most a week in it. In Plymouth, water must be 

 brought from outside the Breakwater, and only this water must be 

 employed for both fertilisation and the subsequent culture. 



At the end of twenty-four to thirty hours, all the eggs which are 

 developing normally will have reached the blastula stage, and have risen 

 to the surface. They must then be decanted off into culture jars. The 

 jars which I used at this stage in the experiments were of a deep bell- 

 shape, and of a capacity of two gallons. Each was fitted with a Browne 

 plunger. This invaluable apparatus has already been described else- 

 where. It is sufficient here to notice that by means of it the water in 

 the jars is kept in constant though gentle agitation, and the formation 

 of a surface film of dust, bacteria, etc., effectually prevented. 



The jars must be carefully guarded from direct sunlight, which soon 

 proves fatal to the larvte. They must be covered in as much as is 

 consistent with the free motion of the plunger, and the water ought to 

 be changed to the extent of one-third of its bulk every day. This is 

 eff'ected by temporarily stopping the action of the plunger, when the 

 healthy larvse will come to the top. The bottom water can then be 



